One of the principal mechanisms by which cellular regulation is effected is through the transduction of extracellular signals across the membrane that in turn modulate biochemical pathways within the cell. Protein phosphorylation, orchestrated by enzymes known as kinases, represents one course by which intracellular signals are propagated from molecule to molecule resulting in a cellular response. These signal transduction cascades are highly regulated and often overlapping as evidenced by the existence of many protein kinases as well as phosphatases, which remove phosphate moieties. It is currently believed that a number of disease states and/or disorders are a result of either aberrant activation or functional mutations in the molecular components of kinase cascades. Consequently, considerable attention has been devoted to the characterization of kinases, especially those involved in energy metabolism. One such kinase is glycogen synthase kinase 3.
Two different mammalian isoforms of glycogen synthase kinase 3 have been identified and each is encoded by a separate gene (Shaw et al., Genome, 1998, 41, 720-727; Woodgett, Embo J., 1990, 9, 2431-2438). These isoforms, designated alpha and beta are expressed in different cell types and in different proportions. In some cells, the expression of these isoforms is under developmental control.
Glycogen synthase kinase 3 beta (also known as tau protein kinase I and GSK3B) is a serine/threonine protein kinase first described as a factor involved in glycogen synthesis. In this pathway, glycogen synthase kinase 3 phosphorylates select residues of glycogen synthase, the rate-limiting enzyme of glycogen deposition, thereby inactivating the enzyme. Therefore, glycogen synthase kinase 3 plays a predominant role in glycogen metabolism and has consequently been investigated as a potential therapeutic target in disease conditions such as diabetes and insulin regulation disorders (Cross et al., FEBS Lett., 1997, 406, 211-215; Eldar-Finkelman et al., Proc. Natl. Acad. Sci. U.S.A., 1996, 93, 10228-10233; Eldar-Finkelman and Krebs, Proc. Natl. Acad. Sci. U.S.A., 1997, 94, 9660-9664; Eldar-Finkelman et al., Diabetes, 1999, 48, 1662-1666). Upstream, glycogen synthase kinase 3 beta is regulated by protein kinase C (Goode et al., J. Biol. Chem., 1992, 267, 16878-16882).
It has been demonstrated that glycogen synthase kinase 3 beta is identical to a previously identified protein known as tau protein kinase-I, which phosphorylates tau, a protein component of paired helical filaments (PHF) found in Alzheimer's brains (Ishiguro et al., FEBS Lett., 1993, 325, 167-172; Lovestone et al., Neuroscience, 1996, 73, 1145-1157; Yamaguchi et al., Acta. Neuropathol. (Berl), 1996, 92, 232-241). The accumulation of these filaments is implicated in the pathological change in brain tissue (Ishiguro et al., FEBS Lett., 1993, 325, 167-172). Glycogen synthase kinase 3 beta is enriched in brain and due to its ability to phosphorylate the tau protein, has been suggested to play a critical role in the development of Alzheimer's disease (Pei et al., J. Neuropathol. Exp. Neurol., 1997, 56, 70-78). Increased synthesis of the enzyme has been shown to increase the cellular maturation of another protein related to Alzheimer's disease, APP (Aplin et al., Neuroreport., 1997, 8, 639-643). It is the aberrant processing of APP that leads to deposition of a beta amyloid in neuritic plaques.
Currently, there are no known therapeutic agents which effectively inhibit the synthesis of glycogen synthase kinase 3 beta and to date, investigative strategies aimed at modulating glycogen synthase kinase 3 beta function have involved the use of antibodies, antisense technology and chemical inhibitors. Disclosed in U.S. Pat. No. 5,837,853 are antisense oligonucleotides targeting the nucleotides which encode the first six amino acids of human glycogen synthase kinase beta intended for use in the treatment of Alzheimer's disease and the prevention of neuronal cell death (Takashima et al., 1998). Disclosed in the PCT publication WO 97/41854 are methods to identify inhibitors of glycogen synthase kinase 3 and the use of these inhibitors for the treatment of bipolar disorders, mania, Alzheimer's disease, diabetes and leukopenia (Klein and Melton, 1997). Other inhibitory compounds are disclosed in WO 99/21859. These heterocyclic compounds are intended for the treatment of a disease mediated by a protein kinase, one of which is glycogen synthase kinase 3 (Cheung et al., 1999). Two other compounds, lithium and valproate, both used in the treatment of bipolar disorders, have been shown to inhibit glycogen synthase kinase 3 beta activity (Chen et al., J. Neurochem., 1999, 72, 1327-1330; Hong et al., J. Biol. Chem., 1997, 272, 25326-25332).
There remains, however, a long felt need for additional agents capable of effectively inhibiting glycogen synthase kinase 3 beta function. The pharmacological modulation of glycogen synthase kinase 3 beta activity or expression may therefore be an appropriate point of therapeutic intervention in pathological conditions.
Antisense technology is emerging as an effective means for reducing the expression of specific gene products and may therefore prove to be uniquely useful in a number of therapeutic, diagnostic, and research applications for the modulation of glycogen synthase kinase 3 beta expression.
The present invention provides compositions and methods for modulating glycogen synthase kinase 3 beta expression.